Method of testing adequacy of cells in a specimen

ABSTRACT

The present invention provides a device and method for determining the adequacy of squamous (ectocervical) cells, columnar (endocervical) cells, neutrophils, and noncellular material in a liquid based cytology specimen. The invention first analyzes a liquid based cytology specimen using light scatter to create a light scatter characteristic representing a predetermined cell. Next the invention determines the presence of squamous (ectocervical) cells versus columnar (endocervical) cells versus neutrophils versus noncellular material using the results of the light scatter. The light scatter characteristic that may be used may be forward light scatter, side light scatter, or both side and forward light scatter.

[0001] This application is a continuation-in-part of U.S. Pat. No.6,329,164, filed Dec. 5, 2000, and granted on Dec. 11, 2001.

BACKGROUND OF THE INVENTION

[0002] The present invention relates to an apparatus and method for usein pre-screening or determining the adequacy of target cells in aspecimen prior to conducting further diagnostic testing or analysis ofthe specimen. More specifically, the present invention concerns a deviceand method which uses light scatter techniques to detect the presence ofa target cell in a specimen.

SUMMARY OF THE INVENTION

[0003] Prior to conducting an analysis or testing of a cell specimen, itis important to insure that an adequate amount of target cells arepresent in the specimen. This is particularly true with respect to papsmear specimens.

[0004] It is estimated that about 180 million pap smears are performedin the United States annually with an estimated 33% of all specimensoriginally collected containing insufficient target cells—ectocervicalor squamous cells. This results in an inability to properly analyze thespecimen and in a tremendous loss in time and money. Typically, not onlymust a patient reschedule another office visit to provide a secondspecimen, a second charge is often incurred to obtain the secondspecimen with no guarantee that enough target cells were again obtained.Thus, there is a need to provide a method and apparatus which provides aquick and efficient system to pre-screen specimens to determine ifadequate target cells are present.

[0005] The present invention solves this lack of pre-screening byproviding a device and method which analyzes a specimen through the useof submitting the specimen to a light scatter analysis. Through thistechnique, a specimen may be analyzed for the presence of sufficientquantities of a target cell. In addition, the technique would also allowa specimen to be analyzed for the presence of target cells which mayadversely affect the diagnostic procedure to be used.

DESCRIPTION OF THE DRAWINGS

[0006] These and other features, objects and advantages of the presentinvention will become apparent from the following description anddrawings wherein like reference numerals represent like elements inseveral views, and in which:

[0007]FIG. 1 shows a sample analysis using light scatter to locate aplurality of predetermined target cells.

DESCRIPTION OF THE PREFERRED EMBODIMENT

[0008] Set forth below is a description of what are currently believedto be the preferred embodiments or best examples of the inventionclaimed. Future and present alternatives and modifications to thepreferred embodiments are contemplated. Any alternates or modificationsin which insubstantial changes in function, in purpose, in structure orin result are intended to be covered by the claims of this patent.

[0009] The present invention may be useful in pre-screening ordetermining the presence of target cells in sufficient quantities priorto subjecting the specimen to further testing. As explained above,screening specimens for adequate amounts of target cells or the presenceof target cells which are adverse to a procedure, while a patient isstill at a medical facility, will save both time and money.

[0010] To do this, the present invention uses a technique commonlyreferred to as light scatter. In this technique, a cell is subjected tolight applied at a predetermined orientation. Because each cell has aspecific morphology, when illuminated, the cell's morphology will causelight to be scattered in a predetermined pattern in both a forward andside direction. Thus, for each target cell of interest, a predeterminedlight scatter pattern can be established in a forward direction, sidedirection or both forward and side directions using flow cytometry andother similar techniques which are known to those of skill in the art.

[0011] Thus, in using the present invention, after a specimen isobtained, it can be quickly subjected to a light scatter technique todetermine if adequate target cells are present for further testingpurposes or if adverse target cells are present. As explained above,using this technique would increase the efficiency andcost-effectiveness of any testing procedure including, but not limitedto, pap smear testing.

[0012] More specifically, to measure the adequacy of liquid basedcervical cytology specimens such as in a pap smear specimen, 1 mL of thesample (usually 10 mL) routinely collected in industry standardpreservatives (Cytyc, TriPath) may be pelleted by centrifugation at500-1000 × g and resuspended in 300 uL of phosphate buffer saline (PBS),pH 7.4. The sample is then run in a flow cytometer or the like foranalysis of forward light scatter and side (90°) light scatter.Ectocervical (columnar) cells, endocervical (squamous) cells,neutrophils, and non-cellular material/debris are resolved into 4cellular populations as shown in FIG. 1 using a log scale rather than alinear scale. The presence of both ectocervical and endocervical cellswould indicate an adequate specimen. Conversely, the absence or the lowamount of a predetermined light scatter pattern for a target cell wouldindicate that an inadequate specimen had been obtained and thecollection procedure should be repeated. This process would takeapproximately 10 minutes.

[0013] In addition, another embodiment of the present invention includesusing a cellular dye such as a nuclear, cytoplasmic, or membrane dye tomeasure the adequacy of a specimen. The cellular dye may be used incombination with the light scatter techniques described above orwithout. For instance, dyes which may be used include, but are notlimited to, nuclear dyes such as propidium iodide or DAPI, cytoplasmicdyes such as eosin, or cell membrane dyes such as DiD, DiO, or DiI.

[0014] The dye may be added to the specimen. Then, in addition to thetechniques described above, the dye is excited by ways known to those ofskill in the art such as by flow cytometry and other similar techniques.This produces in the specimen a multi-parameter pattern for detectingthe target cell of interest. The multi-parameter pattern will be acombination of the flourescent emissions of the dye and light scatter.This technique may be used to determine the adequacy of squamous(ectocervical) cells, columnar (endocervical) cells, neutrophils, andnoncellular material in a liquid based cytology specimen. The targetcells may be ectocervical cells, endocervical cells and other cells thatmay be used in a cervical cancer screening test, as well as otherscreening tests.

[0015] While the preferred embodiments of the present invention havebeen illustrated and described, it will be understood by those ofordinary skill in the art that changes and other modifications can bemade without departing from the invention in its broader aspects.Various features of the present invention are set forth in the followingclaims.

What is claimed is:
 1. A method of determining the adequacy of squamous(ectocervical) cells, columnar (endocervical) cells, neutrophils, andnoncellular material in a liquid based cytology specimen comprising: a.adding a dye to said specimen; b. analyzing said liquid based cytologyspecimen using flourescent emission and light scatter to create acharacteristic multi-parameter pattern representing a predeterminedcell; and c determining the presence of squamous (ectocervical) cellsversus columnar (endocervical) cells versus neutrophils versusnoncellular material using the results of said multi-parameter patternanalysis.
 2. The method of claim 1 wherein said light scatter is forwardlight scatter.
 3. The method of claim 1 wherein said light scatter isside light scatter.
 4. The method of claim 1 wherein said light scatteris both side and forward light scatter.
 5. A method of determining theadequacy of at least one target cell in a specimen comprising: adding adye to said specimen; using flourescent emission and light scatter todetermine if an adequate amount of at least one target cell is presentin the specimen prior to subjecting the specimen to further analysis. 6.The method of claim 5 wherein the target cell is an ectocervical cell.7. The method of claim 5 wherein the target cell is an endocervicalcell.
 8. The method of claim 5 wherein the specimen is to be used in acervical cancer screening test.
 9. The method of claim 5 wherein saidlight scatter is forward light scatter.
 10. The method of claim 5wherein said light scatter is side light scatter.
 11. The method ofclaim 5 wherein said light scatter is both side and forward lightscatter.
 12. A method of determining the adequacy of at least one targetcell in a pap smear specimen comprising: adding a dye to said specimen;using flourescent emission and light scatter to determine if an adequateamount of at least one target cell is present in the specimen prior tosubjecting the specimen to further analysis.
 13. The method of claim 12wherein the target cell is an ectocervical cell.
 14. The method of claim12 wherein the target cell is an endocervical cell.
 15. The method ofclaim 12 wherein said light scatter is forward light scatter.
 16. Themethod of claim 12 wherein said light scatter is side light scatter. 17.The method of claim 12 wherein said light scatter is both side andforward light scatter.